Difference between revisions of "Part:BBa K1088013"

Revision as of 21:25, 25 September 2013

B. subtilis dxs (lac promoter with lac inhibitor: IPTG inducible)

This part consist of the dxs gene derived from B. subtilis under the control of the lac promoter and has a strong RBS.

To repress expression from the lac promoter, the lacI:LVA gene (under a constitutive promoter, with a strong RBS and a efficient terminator) was placed counter-clockwise to the dxs gene. The repression can be relieved with addition of IPTG, which binds and inhibits the function of LacI:LVA.

The levels of expression was meassured with a similar Brick with linker-GFP fused to Dxs (BBa_1088009) in E. coli K-12 MG1655 using fluorescence activated cell sorting (FACS). The experiment proved that the reporter fusion was only very weakly expressed when the strain was grown without IPTG in the media, and that the reporter fusion was expressed at much higher levels after addition of IPTG. Though, in the same experiment we compared the LVA-tagged LacI with the natural and found that the natural LacI responded better to the IPTG addition. See experience for more details.

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal EcoRI site found at 2505
Illegal EcoRI site found at 3162
Illegal PstI site found at 2563
Illegal PstI site found at 3009
12
INCOMPATIBLE WITH RFC[12]
Illegal EcoRI site found at 2505
Illegal EcoRI site found at 3162
Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal PstI site found at 2563
Illegal PstI site found at 3009
21
INCOMPATIBLE WITH RFC[21]
Illegal EcoRI site found at 2505
Illegal EcoRI site found at 3162
23
INCOMPATIBLE WITH RFC[23]
Illegal EcoRI site found at 2505
Illegal EcoRI site found at 3162
Illegal PstI site found at 2563
Illegal PstI site found at 3009
25
INCOMPATIBLE WITH RFC[25]
Illegal EcoRI site found at 2505
Illegal EcoRI site found at 3162
Illegal PstI site found at 2563
Illegal PstI site found at 3009
Illegal NgoMIV site found at 2462
Illegal AgeI site found at 2355
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 2304
Illegal SapI.rc site found at 3003

@@ Line 6: / Line 6: @@
 To repress expression from the lac promoter, the ''lacI:LVA'' gene (under a constitutive promoter, with a strong RBS and a efficient terminator) was placed counter-clockwise to the ''dxs'' gene. The repression can be relieved with addition of IPTG, which binds and inhibits the function of LacI:LVA.
-The levels of expression was meassured with a similar Brick with linker-GFP fused to Dxs (BBa_1088009) in ''E. coli'' K-12 MG1655 using fluorescence activated cell sorting (FACS). The experiments proved that there was low expression when grown without IPTG and that there was high expression after addition of IPTG. Though, in the same experiment we compared the LVA-tagged LacI with the natural and found that the natural LacI responded better to the IPTG addition. See experience for more details.
+The levels of expression was meassured with a similar Brick with linker-GFP fused to Dxs (BBa_1088009) in ''E. coli'' K-12 MG1655 using fluorescence activated cell sorting (FACS). The experiment proved that the reporter fusion was only very weakly expressed when the strain was grown without IPTG in the media, and that the reporter fusion was expressed at much higher levels after addition of IPTG. Though, in the same experiment we compared the LVA-tagged LacI with the natural and found that the natural LacI responded better to the IPTG addition. See experience for more details.
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